Abstract Colorectal cancer (CRC) remains the third most common cause of cancer-related deaths in the U.S. The majority of CRC tumors are non-immunogenic, i.e. they lack a significant intensity of anti-tumor immune response, and are typically unresponsive to immunotherapies that have dramatically changed the way we treat many cancer patients. Several chemokines and cytokines are implicated in CRC immune response. One of the under examined features of CRC is the regulation of anti-tumor immunity by atypical chemokine receptor 4 (ACKR4) expression. This receptor, is involved in the internalization and degradation of multiple chemokines, such as chemokine (C-C motif) ligand 19 (CCL19) and CCL21, thereby modulating the CC-chemokine receptor 7 (CCR7)/CCL19/CCL21 chemotaxis and its downstream immune responses. However, the role of ACKR4 in immune regulation? including the trafficking and differentiation of immune cells in a CRC environment?remains to be investigated. Our preliminary data showed that overexpression of ACKR4 is correlated with high CD8+ T-cell infiltration and with the overall immune response (indicated by the ?immunoscore?) in CRC patients' samples. Therefore, we hypothesize that ACKR4 positively affects immune regulation in CRC and confers antitumor immunity by modulating CCR7/CCL19/CCL21 chemotaxis. In Aim 1, we will establish the connection between ACKR4 expression and anti-tumor immunity of CRC. We will generate orthotopic preclinical models mouse colorectal cancer different levels of ACKR4 expression levels via small animal endoscopy and microinjection. Tumor volume, numbers of CD8+ T cell and NK cells, and key cytotoxic cytokines will be measured. We will also determine the primary immune cells that respond to ACKR4 expression in CRC tumor models. Aim 2 will decipher the mechanisms and regulation of antitumor immunity by ACKR4 in CRC. Since ACKR4 is the scavenging receptor of CCL19 and CCL21, we will focus on the CCR7/CCL19/CCL21 chemotaxis axis. Results will be further confirmed using CCL21-/-, CCL19-/- as the host mice for orthotopic CRC models. Clinical Impact. The results from this pilot study will establish the clinical significance of ACKR4 expression levels to immune response in colorectal cancer. As an outcome, it will fundamentally advance our knowledge of how anti-tumor immunity was generated and regulated in CRC, and provide novel targets for CRC immunotherapies.